Cellular senescence was first described by Hayflick and Moorhead (1961) when they observed that normal human fibroblasts entered a state of irreversible growth arrest after serial passage in vitro. In contrast, cancer cells did not enter this growth arrested state and proliferated indefinitely. The maximum number of cell divisions that a cell can undergo, termed the Hayflick limit, varies from cell type to cell type and organism. In fibroblasts, this number is about 50 divisions, after which cell division ceases.
The process of cellular senescence can be triggered by multiple mechanisms, including telomere shortening, derepression of the INK4a/ARF locus, and DNA damage. As discussed below, all three of these mechanisms implicate the function of the tumor suppressor protein p53.
Telomere shortening provides a mechanism capable of counting cell divisions. Telomeres consist of repetitive DNA elements at the end of linear chromosomes that protect chromosome ends from degradation and recombination. Due to the intrinsic inability of the DNA replication machinery to copy the ends of linear molecules, telomeres become progressively shorter with each round of replication, thus providing a counting mechanism for keeping track of the number of cell divisions that have occurred in a population of cells. As increasing numbers of cell division occur, the telomeres reach a critically short length, which present as double-stranded DNA breaks that activate the p53 tumor suppressor protein resulting in telomere-initiated senescence or apoptosis.
Derepression of the INK4a/ARF locus can also serve as a cell division counting mechanism. The INK4a/ARF locus is normally expressed at very low levels in most tissues of young organisms but progressively becomes derepressed with aging. Thus, a cell division counting mechanism is provided by a progressively increased level of repression of the INK4a/ARF locus. The p16INK4a protein functions as an inhibitor of cyclin-dependent kinases CDK4 and CDK6, thus providing a G1 cell cycle arrest. ARF regulates p53 stability through inactivation of the p53-degrading ubiquitin ligase MDM2.
The accumulation of DNA damage over time can also serve as a trigger for cell senescence. As an organism ages, increases in DNA mutations, DNA oxidation, and chromosome losses are observed. These observations have prompted investigators to consider DNA damage as contributing to cellular senescence and organismal aging. As a guardian of cell cycle progression after DNA damage, p53 plays a role here too, as p53 induces the expression of the cell cycle inhibitor p21 when a cell has undergone DNA damage.
Given the direct impact that cell senescence has on cell division and cell cycle arrest, one would expect this process to play a central role in such diverse processes as aging, cancer, and tissue regeneration. The present invention provides methods and compositions for manipulating these diverse processes through the modulation of cell senescence.